中波紫外线对体外培养皮肤干细胞某些标记分子表达的影响

Changes of molecular markers in cultured skin stem cells exposed to ultraviolet B （UVB） in vitro

目的 探讨中波紫外线（UVB）对体外培养皮肤干细胞某些标记分子表达的影响.方法 利用快速贴壁法分离、培养皮肤干细胞,K15、β-连环素鉴定皮肤干细胞.利用峰值在305 nm的UVB照射皮肤干细胞,照射剂量为10 mJ/cm2.用免疫组化法分别检测经照射组与对照组皮肤干细胞CD34、β-连环素、p53的表达变化.结果 未经UVB照射的皮肤干细胞密度大,细胞呈圆形或多角形,细胞形态清晰,胞质均匀,可见有核分裂细胞,核质比例大.β-连环素主要在细胞膜和细胞质表达,其胞膜和胞核染色阳性率分别为64.74%和8.4%;p53主要在胞质表达,其胞核染色阳性率为6.9%.UVB照射后的细胞密度稀,细胞变形、不规则,胞质出现空泡,核质比例变小,部分细胞出现核固缩和凋亡.β-连环素主要在细胞质和细胞核表达,其胞膜和胞核染色阳性率分别为64.74%和0;p53主要在细胞核表达,其胞核染色阳性率为100%.CD34在两种情况下均不表达.结论 UVB能使CD34阴性皮肤干细胞β-连环素在细胞质聚集并移入细胞核,p53可定位于皮肤干细胞的细胞质,经UVB照射后可移入细胞核.

Objective To investigate the changes of molecular markers in cultured skin stem cells exposed to UVB in vitro. Methods Skin stem cells were isolated and cultured according to their adherasion ability,and identified by immunohistochemistry using anti-K15 and anti-β-integrin antibodies. Then, a part of the skin stem cells were irradiated with UVB at 10 mJ/cm2 for 2 times. After 24-hour additional culture, the expressions of CD34, beta-catenin and p53 were detected with immunohistochemistry. Results Skin stem cells showed a high density in culture free of irradiation, which were round or polygon with a clear shape, well-distributed cytoplasm, high N/C ratio; mitotic cells could be seen. In unirradiated skin stem cells, beta-catenin was expressed predominantly in cell membrane and cytoplasm, with a positive expression rate of 64.74% and 8.4%in membrane and cytoplasm respectively; p53 was expressed mainly in cell cytoplasm and nuclei, with a positive expression rate of 6.9% in cell nuclei. After exposure to UVB, skin stem cells decreased in cell density and N/C ratios with a deformed and anomalous shape, vacuoles were present in cytoplasm, and some cells experienced karyopyknosis or apoptosis. Additionally, in irradiated cells, beta-catenin was expressed predominantly in cytoplasm with a positive expression rate of 64.74% and 0 in cytoplasm and nuclei, respectively; p53 was expressed mainly in nuclei with a positive expression rate of 100%. CD34 was detected in neither unirradiated nor irradiated skin stem cells. Conclusion UVB can promote beta-eatenin to accumulate in cytoplasm as well as beta-catenin and p53 to migrate from cytoplasm to nuclei.